Ebola virus (EBOV) is a negative single-stranded RNA virus of the filovirus family that is listed as a NIAID Category A agent. The long-term aim of this project is to develop cytomegalovirus (CMV)-based vaccines to interfere with EBOV transmission in wild Great Ape populations. Human outbreaks of severe EBOV-associated hemorrhagic fever in Western Africa have been increasing in recent years, resulting in high morbidity and mortality with a death rate approaching 90%1. The natural reservoir and mode of transmission to humans is not defined, however bats and Great Apes (gorillas and chimpanzees) appear to play a role in transmission. None of the vaccine candidates that have been shown to protect against EBOV in rodent and non-human primate (NHP) models have the ability to spread efficiently from initial vaccine recipients to other members of the population1,7-16. Thus, population immunization requires that most individuals are vaccinated. This inability to disseminate poses difficulties when targeting EBOV vaccines to inaccessible Western African Great Ape populations. A vaccine using CMV-based vectors expressing EBOV antigens may be ideal for such inaccessible wild populations. CMV possesses the remarkable ability to re-infect and disseminate through the population regardless of prior CMV immunity17-19. CMV-based vaccines are also highly immunogenic17,20-24. In our strategy, vaccination of 'founders'would initially spread the vaccine through the population, eliminating the need for immunization of all individuals. CMV is also transmitted vertically, thus a CMV-based vaccine could afford a permanent solution to preventing EBOV transmission. CMV is most likely ubiquitous in wild gorillas and chimpanzees25-27. A CMV-based vaccine merely represents infection with a benign, ubiquitous virus, differing only by expression of EBOV-derived antigens. EBOV exists as five antigenically distinct species. We propose to construct CMV-based vectors individually expressing EBOV glycoprotein (GP) (a major, but highly variable target of the humoral immune response) and EBOV nucleoprotein (NP) (a more highly conserved protein containing multiple T cell epitopes). Targeting both GP and NP may induce heterologous (cross-EBOV species) as well as homologous protection. This initial study to establish the principle of a founder-based vaccine for EBOV is ideally suited to R21 funding. Establishment of 'proof-of-principle'with MCMV in mice will support an R01 application to assess protection afforded by GP and NP-expressing rhesus CMV-based vectors in the NHP model, wherein levels of cross-protection can also be assessed. Specific Aim 1: will construct and characterize in vitro two murine CMV (MCMV) vector constructs individually expressing either EBOV GP or NP (designated MCMV/EBOV-GP and -NP, respectively). Specific Aim 2: will determine the ability of MCMV/EBOV vectors to induce and maintain an EBOV GP and NP-specific humoral and T cell response in mice. Specific Aim 3: will determine the capacity of MCMV/EBOV vectors to protect against mouse-adapted EBOV challenge in a mouse model. PUBLIC HEALTH RELEVANCE: Ebola virus (EBOV) is a NIAID Category A agent that causes outbreaks of lethal haemorrhagic fever in endemic areas of Western Africa. The frequency of outbreaks has been on the rise, which combined with the potential for accidental and deliberate release into non-endemic nations, makes EBOV a major health concern. The long-term aim of this project is to develop cytomegalovirus (CMV)-based vaccines to interfere with EBOV transmission in wild Great Ape populations, and thereby block this infectious conduit for EBOV transmission to Man.